SL0439: A structure-tuned bispecific nanobody CAR-T cells targeting BCMA/GPRC5D for relapsed/refractory multiple myeloma with extramedullary disease

Abstract Background: Despite B-cell maturation antigen (BCMA)-directed chimeric antigen receptor (CAR) T-cell therapies have demonstrated remarkable efficacy in relapsed or refractory multiple myeloma (r/r MM), single-target approaches carry risks of immune evasion by BCMA-negative clones during prolonged treatment, potentially limiting long-term survival benefits. To address this, we developed structure-optimized BCMA/GPRC5D bispecific nanobody-based CAR-T cells to enhance antitumor efficacy in r/r MM patients. Aims: We aim to develop and validate a structure-optimized, bispecific nanobody-based BCMA/GPRC5D CAR-T construct through comprehensive in vitro screening. Additionally, we conduct a phase Ⅰ clinical trial (NCT07003568) to evaluate its efficacy and safety in RRMM patients. Methods: We developed a bispecific BCMA/GPRC5D-targeting CAR incorporating optimized nanobodies and flexible linkers within a second-generation 4-1BB-containing CAR framework to ensure stable dual-target binding. The lead construct was tested in a single-arm, multicenter phase Ⅰ trial (BOIN design) in RRMM patients (aged 18–75, ECOG 0–3). The starting dose of SL0349 CAR-T cells was 1.0×10⁶ cells/kg. Throughout the trial, we assessed clinical responses, adverse events, and CAR-T cell kinetics of SL0349. Results: Through alpaca immunization (6 rounds) with BCMA or GPRC5D antigens and phage display screening, we identified 6 BCMA- and 6 GPRC5D-specific nanobodies for CAR construction.The in vitro experimental screening demonstrated that the optimized bispecific CAR candidate SL0349 exhibited significantly enhanced cytotoxic activity against double-positive tumor cells when compared to monospecific CAR constructs and possessed superior T-cell transduction characteristics. Notably, SL0349 displayed superior expansion capacity, stable CAR expression, enhanced cytokine production, and sustained antitumor efficacy following repeated antigen exposure, in comparison to other control constructs.Between May 17, 2025, and July 17, 2025, 5 patients were enrolled and received CAR-T products at Shanghai Liquan Hospital.2 were male and 3 were female patients, with 2 was white and 3 were Asian respectively. Median age was 55.2 years (range,47-60). Median line of previous therapies was 3 (range,2-4) and median time since diagnosis was 57 months (range,14-81). Four patients had Revised International Staging System II or III disease, 4 patients had a history of auto-HSCT, and 1 patient had a history of allo-HSCT. 4 patients had high-risk cytogenetic abnormalities, and 4 patients had extramedullary diseases(EMDs). All patients received 1e6/kg CAR-T cells. No DLTs were observed. The most common grade 3 or worse adverse events were hematological toxicities in 5 patients, including 3 (60%) patients with neutropenia, 4 patients with anemia, and 4 (100%) patients with thrombocytopenia. Two patients had severe pancytopenia before CAR T treatment. All patients had cytokine release syndrome (CRS) with Grade 1(80%) and Grade 3(20%). No Immunal neurotoxicity were observed. One patient received continuous hemofiltration therapy due to tumor lysis syndrome complicated with acute renal failure. Two patients had Grade 2 pulmonary fungal infection, another had Grade 3 soft tissue infection, and one developed Grade 3 acute cholecystitis. One patient who underwent alloHSCT developed Grade 3 aGVHD (skin, liver, and intestine). By the cutoff date (July 16, 2025), the overall response rate was 80% in the entire cohort, including 2 (40%) Stringent complete response, sCR), one (25%) very good partial response (VGPRs), one (25%) partial response (PR) and one (25%) stable disease (SD). The median time to best response was 21 days (14-28) after CAR T-cells infusion. Four patients obtained measurable residual disease (MRD) negativity and two patients had got CR results for EMDs. CAR T cell's Peak expansion (Cmax) was 565510 (range:8430-1107380) copies /μg DNA with a median Tmax of 16.5 days (range 10-26d). Conclusion: The BCMA/GPRC5D bispecific nanobody-based CAR-T (SL0349) has demonstrated superior structural properties and shows promising potential as a safe and effective therapeutic option for RRMM patients. Notably, it exhibits unique capabilities in eliminating EMDs and reducing the risk of antigen-negative relapse post CAR-T infusion. Disclosures：CAR vector is provided by Heibei Senlang Biotechnology Co., Ltd. CAR-T manufacturing and QC control are conducted in Beijing Gobroad Hospital.